Botulinum neurotoxins (BoNTs) are extremely toxic proteins and can be classified into distinct subgroups based, inter alia, on peptide sequence and substrate specificity. All of the naturally occurring BoNTs (BoNT/A-G) are composed of a heavy chain that mediates toxin entry into a target cell and a light chain with zinc-dependent protease activity that hydrolyzes selected SNARE proteins that mediate fusion of neurotransmitter vesicles to the membrane that forms part of the synaptic cleft. For example, the light chain of BoNT/A hydrolyzes with high specificity SNAP-25, which is required for vesicle-mediated exocytosis of acetylcholine into the synaptic cleft.
Unfortunately, therapeutic options for BoNT poisoning (e.g., immunoprophylactic vaccine, antitoxin or antibody administration, etc.) have proven less than ideal for various reasons. For example, vaccination typically results in relatively low antibody titers, while neutralizing proteins are often immunogenic. Still further, the therapeutic window of such options is generally limited to a small period as the BoNT is readily taken up into the neural cells. Therefore, development of various small molecule inhibitors of the light chain protease activity has gained considerable attention.
For example, various small-molecule compounds have recently been reported as light chain protease inhibitor, and most notably selected compounds that include a hydroxamate group (see e.g., Bioorg Med Chem Lett. 2007 December 1; 17(23): 6463-6466). However, at least some of these compounds are relatively toxic and thus failed to provide a meaningful antidote. Similarly, various quinolinol compounds were reported as relatively active protease inhibitors (see e.g., Antimicrobial Agents And Chemotherapy, August 2009, p. 3478-3486). However, such compounds tend to suffer from relatively poor solubility. In still further known approaches, the small molecule inhibitor NSC 240898 was designed based on docking data from pseudo-peptide mpp-RATKML, which is a known inhibitor of BoNT/A (U.S. Pat. App. No. US2007/0112049 or The Journal Of Biological Chemistry Vol. 282, NO. 7, pp. 5004-5014). While such approach provided at least a promising scaffold, the affinity of the NSC 240898 is less than desirable. Further screening data and lead compounds are provided elsewhere (see e.g., Proc Natl Acad Sci USA. 2007 February 20; 104(8): 2602-2607; J Appl Toxicol. 1999 December; 19 Suppl 1:S5-S11; U.S. Pat. No. 7,574,340), however, most of the known compounds are either toxic or have a relatively weak activity.
As can be seen from the foregoing, there is a relatively wide diversity in structures that have at least some inhibitory activity against BoNT protease activity. However, all or almost all of the known compounds suffer from one or more disadvantages. Therefore, there is a continuing need for new and improved methods and compositions for inhibiting BoNT related protease activity.